Cloning and nucleotide sequence of a lipase gene from a soil isolate

Abstract

The present study was aimed to isolate and characterize the lipolytic enzyme producing bacteria from soil samples of regions around Zayande-rood river of Isfahan, Iran. Soil samples were collected from 15 cm depth of soil surface. Based on morphology, distinct colonies were isolated and purified through streak culture on to standard agar plates. Isolated colonies were examined for lipase activity using egg-yolk agar medium. Total of 15 isolates developed clear zones around their growth area which were considered as lipase positive. Preliminary identification of lipolytic active isolates revealed a gram-positive, rod-shaped, endospore-forming and catalase positive bacteria, characteristics indicative of the genus bacillus. The gene coding for an extracellular lipase was cloned using PCR techniques. The gene was identified to be 639 bp in length and encoded a peptide of 212 amino acids with calculated molecular mass of 19353 Da, and pI 9.28. The DNA sequence and deduced amino acid sequence of the hypothetical lipase showed striking similarities to lipases from B. subtilis strains.